Staphylococcus aureus is a facultative anaerobic, Gram-positive bacterium discovered by Dr. Alexander Ogston in 1880. Literature reports suggest that about 30% to 50% of the population has been carriers of S. aureus at one time in their lives and about 20% are long-term carriers. S. aureus is widespread in the environment and has become one of the most commonly isolated pathogens in hospital-acquired infections. Moreover, S. aureus can cause numerous illnesses, from minor skin infections to life-threatening diseases, such as abscesses, pneumonia, meningitis, endocarditis, and septicemia. According to reports of the National Institutes of Health and Centers for Disease Control and Prevention, S. aureus infects 500,000 people yearly in America, more than 94,000 of which are cases of life-threatening, antibiotic-resistant S. aureus infections.
Bacterial culture and metabolic tests are standard protocols for bacterial identification in use by most hospitals. But this process might take days for identification of the pathogenic bacteria—an unacceptable delay in emergency and critically ill situations such as sepsis. For this reason, several ultrasensitive detection methods based on nucleic acid amplification, such as PCR (polymerase chain reaction), LCR (ligase chain reaction) and SDA (strand displacement amplification), among others, have been introduced. All of these technologies are capable of detecting low numbers of bacterial cells within several hours. However, these technologies require prior isolation of bacterial DNA, preparation of enzyme reaction mix, and expensive instruments for nucleic acid amplification. These high costs and complex procedures limit the widespread use of these technologies for clinical diagnosis. Antibody-based immunoassays for bacterial identification are well established and have been used for many years. However, ultrasensitive detection with such approaches is limited by the fact that antibodies are proteins and thus cannot be amplified. This limitation was circumvented by the development of a technology called immuno-PCR, in which the antibody is cross-linked with a DNA “barcode” for PCR amplification. Although this technology is sensitive, the conjugation and purification of antibody-DNA complexes is still a daunting task.
Aptamers are DNA or RNA molecules that can fold into a variety of structures. Like an antibody, a good aptamer can specifically bind to its target with pico- to nanomolar affinity. Importantly, unlike antibodies, aptamers can be directly amplified by PCR. Since their discovery in the late 1990s, aptamers have been widely used in many applications, including target detection, enzyme inhibition, receptor regulation, and drug delivery. Several bacterial aptamers had been isolated and recently used in identification of bacteria, including Escherichia coli, Mycobacterium tuberculosis, Salmonella enterica, and Bacillus anthracis. However, none of these studies reported showed the capability of identifying extremely low numbers of target bacteria without a PCR reaction.
Therefore, a heretofore unaddressed need exists in the art to address the aforementioned deficiencies and inadequacies, especially in connection with detection of a biological target present in a low concentration in a sample.